Plasminogen activator inhibitor type 1 (PAI-1) is involved in the regulation of the blood fibrinolytic system. Increased plasma levels of PAI-1 are implicated in the impairment of fibrinolytic function and may be associated with thrombotic diseases. Levels of PAI-1 increase with age and are elevated in conditions such as normal pregnancy and sepsis. The sensitive quantitative measurement of functionally active human PAI-1 in plasma samples is easily performed with this 96 well strip format ELISA kit. The PAI-1 concentration of normal platelet-free plasma is 21 ng/ml, platelet-rich plasma is 283 ng/ml and serum is 270 ng/ml. 1 PAI-1 unit = 1.34 ng. The assay measures active PAI-1 in the 0.125-100 U/ml range. Samples giving human PAI-1 levels above 100 U/ml should be diluted in PAI-1 depleted plasma before use. Normal plasma should be applied directly to the plate for best results. It is important to ensure a platelet free preparation of plasma as platelets can release PAI-1. Functionally active PAI-1 reacts with urokinase coated onto a micro titer plate.
Latent or complexed PAI-1 will not bind to the plate and will not be detected by the assay. Validated in citrate, EDTA, and heparin plasma. Vitronectin does not interfere with the detection of active PAI-1. This kit does not cross react with mouse PAI-1. After appropriate washing steps, anti human PAI-1 primary antibody binds to the PAI-1. Excess antibody is washed away, and bound antibody is reacted with the secondary antibody conjugated to HRP. Following an additional washing step, TMB substrate is used for color development at 450 nm. Color development is proportional to the concentration of PAI-1 in the samples. A standard calibration curve is prepared in PAI-1 depleted plasma using dilutions of purified PAI-1 and is measured along with the test samples. All reagents and standards are provided in these ELISA kits.
The Human PAI-1 standard provided in this kit is calibrated against the WHO International Standard for Human PAI-1 (NIBSC Code 92/654).This assay uses an exclusive enzyme capture technology to only detect functionally active protein.
The standard curve for this kit is generated using included PAI-1 depleted human plasma and is best suited for measurement of PAI-1 in plasma or serum. This kit is also available with a standard curve in BSA/TBS for researchers measuring PAI-1 in culture media or tissue extracts under catalog number HPAIKT-NP.
Suggested additional reagents: 10X Wash Buffer, TMB Substrate, uPA Plate, Secondary Antibody
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